The Redox Potential of the Plastoquinone Pool of the Cyanobacterium Synechocystis Species Strain PCC 6803 Is under Strict Homeostatic Control

A method is presented for rapid extraction of the total plastoquinone (PQ) pool from Synechocystis sp. strain PCC 6803 cells that preserves the in vivo plastoquinol (PQH(2)) to -PQ ratio. Cells were rapidly transferred into ice-cold organic solvent for instantaneous extraction of the cellular PQ plus PQH(2) content. After high-performance liquid chromatography fractionation of the organic phase extract, the PQH(2) content was quantitatively determined via its fluorescence emission at 330 nm. The in-cell PQH(2)-PQ ratio then followed from comparison of the PQH(2) signal in samples as collected and in an identical sample after complete reduction with sodium borohydride. Prior to PQH(2) extraction, cells from steady-state chemostat cultures were exposed to a wide range of physiological conditions, including high/low availability of inorganic carbon, and various actinic illumination conditions. Well-characterized electron-transfer inhibitors were used to generate a reduced or an oxidized PQ pool for reference. The in vivo redox state of the PQ pool was correlated with the results of pulse-amplitude modulation-based chlorophyll a fluorescence emission measurements, oxygen exchange rates, and 77 K fluorescence emission spectra. Our results show that the redox state of the PQ pool of Synechocystis sp. strain PCC 6803 is subject to strict homeostatic control (i.e. regulated between narrow limits), in contrast to the more dynamic chlorophyll a fluorescence signal.